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. 1980 Jul;77(7):4160–4164. doi: 10.1073/pnas.77.7.4160

The four cytoplasmically made subunits of yeast mitochondrial cytochrome c oxidase are synthesized individually and not as a polyprotein.

K Mihara, G Blobel
PMCID: PMC349790  PMID: 6254013

Abstract

Subunit-specific antisera prepared against each of the four cytoplasmically made subunits (IV, V, VI, and VII) of yeast mitochondrial cytochrome c oxidase (EC 1.9.3.1) were used to precipitate immunoreactive polypeptides that were synthesized either in vitro, in a cell-free protein-synthesizing system programmed with total yeast mRNA, or in vivo, in intact cells and in spheroplasts, under conditions of pulse labeling, pulse-chase labeling, and continuous labeling. Using N-formyl-[35S]Met-rTNA as the only radioactively labeled component in the cell-free system, we demonstrated (i) that each of the four cytoplasmically made subunits is synthesized as a separate entity and not as part of a polyprotein as was claimed by others; (ii) that subunits IV, V, and VI are synthesized as precursors, larger by 1500-3000 daltons than their mature counterparts; in contrast, subunit VII is not synthesized as a larger precursor. Precursor forms of subunits IV, V, and VI identical to those synthesized in vitro were also detected in vivo by pulse-labeling of spheroplasts. The observed disappearance of these larger forms after a chase is compatible with the notion that they represent short-lived precursors that are rapidly converted to their mature counterparts during or shortly after import into mitochondria. Furthermore, using N-formyl-[35S]Met-tRNA, we provide definitive evidence that two of the cytoplasmically made subunits (beta and gamma) of another oligomeric inner mitochondrial membrane protein (F1-ATPase, EC 3.6.1.3) are not synthesized as part of a polyprotein but as individual precursors.

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Selected References

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